Male behavior through the breeding season in Saimiri boliviensis boliviensis

The Bolivian squirrel monkey (Saimiri boliviensis boliviensis) is a seasonal breeder. Male squirrel monkeys show distinct morphological and behavioral changes prior to and during the breeding season. A “fatting syndrome” includes increased body weight, increased levels of androgens, and in the Bolivian subspecies, an increasingly active role in the social organization of the group. In this study, the behavior of ten adult male Bolivian squirrel monkeys was analyzed over a 6-month period prior to, during, and after the breeding season. Each was housed as the only adult male in a breeding unit with six to ten adult females and one juvenile male. Employing a principle components method, 11 behavioral clusters were generated from 27 responses. Their activity clusters were identified as follows: sexual activity that showed a peak around the time of peak conceptions; excitatory activity that was initially high but decreased throughout the breeding season; and maintenance activity that did not change across the breeding season. The changing social behavior of the male squirrel monkey parallels physiological changes and is correlated with changing androgen levels.     

Seasonal changes of serum concentrations of estradiol and progesterone in Bolivian squirrel monkeys (Saimiri sciureus)

Serial measurements of estradiol (E2) and progesterone (P) were used to describe the ovarian cycle of the Bolivian squirrel monkey. Group-caged, sexually mature female squirrel monkeys, housed with males, were sampled daily between 0900 and 1100 hr. Sampling was carried out during the breeding and nonbreeding seasons, for periods of 19–20 days from September 1981 to May 1982. Seasonal differences in serum concentrations of E2 and P were found with low levels of E2 and P and an absence of preovulatory surges of E2 during the nonbreeding season. This pattern was also observed in some animals during the breeding season. An abrupt increase in serum P concentrations in December appeared to signal the onset of cycling. Cycling animals had well-defined peaks of E2 (450–9,500 pg/ml) followed by increasing levels of P, which were >200 ng/ml in some animals. After the breeding season, E2 and P levels returned to their initially low levels. Levels of both steroids in cycling animals were higher than those reported for other primates and for previous measurements made in squirrel monkeys. Cycle length based on time interval between consecutive E2 peaks varied from 6–12 days.     

Seasonal Expression of Androgen Receptor,Aromatase, and Estrogen Receptor Alpha and Beta in the Testis of the Wild Ground Squirrel (Citellus Dauricus Brandt)

The aim of this study was to investigate the seasonal expression of androgen receptor (AR), estrogen receptors α and β (ERα and ERβ) and aromatase cytochrome P450 (P450arom) mRNA and protein by real-time PCR and immunohistochemistry in the wild ground squirrel (WGS) testes. Histologically, all types of spermatogenic cells including mature spermatozoa were identified in the breeding season (April), while spermatogonia and primary spermatocytes were observed in the nonbreeding season (June), and spermatogonia, primary spermatocytes and secondary spermatocytes were found in pre-hibernation (September). AR was present in Leydig cells, peritubular myoid cells and Sertoli cells in the breeding season and pre-hibernation with more intense staining in the breeding season, whereas AR was only found in Leydig cells in the nonbreeding season; P450arom was expressed in Leydig cells, Sertoli cells and germ cells during the breeding season, whereas P450arom was found in Leydig cells and Sertoli cells during pre-hibernation, but P450arom was not present in the nonbreeding season; Stronger immunohistochemical signal for ERα was present in Sertoli cells and Leydig cells during the breeding season; ERβ was only expressed in Leydig cells of the breeding season. Consistent with the immunohistochemical results, the mean mRNA level of AR, P450arom, ERα and ERβ were higher in the testes of the breeding season when compared to pre-hibernation and the nonbreeding season. These results suggested that the seasonal changes in spermatogenesis and testicular recrudescence and regression process in WGSs might be correlated with expression levels of AR, P450arom and ERs, and that estrogen and androgen may play an important autocrine/paracrine role to regulate seasonal testicular function.Key words: Wild ground squirrels, testes, seasonal expression, androgen and estrogen receptors, aromatase cytochrome P450, Citellus dauricus Brandt     

Ovarian stimulation of squirrel monkeys (Saimiri boliviensis boliviensis) using pregnant mare serum gonadotropin

The application of assisted reproductive technologies (ART) to nonhuman primates has created opportunities for improving reproductive management in breeding colonies, and for creation of new animal models by genetic modification. One impediment to the application of ART in Saimiri spp. has been the lack of an effective gonadotropin preparation for ovarian stimulation. Pregnant mare serum gonadotropin (PMSG) is inexpensive and readily available, but its repeated use in rhesus monkeys has been associated with induction of a refractory state. We have compared PMSG to recombinant human follicle stimulating hormone (rhFSH) for controlled ovarian stimulation in Bolivian squirrel monkeys. Groups of mature squirrel monkeys received rhFSH (75 IU daily) or PMSG (250 IU twice daily) by subcutaneous injection for 4 d during the breeding season (November to January) or nonbreeding season (March to September). Serum estradiol (E2) was measured daily. Follicular growth was monitored by abdominal ultrasound. During the breeding season, PMSG induced a higher E2 response than did rhFSH, with mean E2 levels being significantly higher within 3 d of stimulation. Superior follicular development in PMSG animals was confirmed by abdominal ultrasonography. During the nonbreeding season, PMSG elicited a similar increase in serum E2 levels despite the fact that basal serum E2 is typically low during the nonbreeding season. Repeated use of PMSG (< or = 3 cycles of administration) produced no attenuation of the E2 response. We conclude that PMSG is highly effective for repeated cycles of controlled ovulation stimulation in the squirrel monkey.     

Seasonal changes in serum dehydroepiandrosterone,androstenedione, and testosterone levels in the squirrel monkey (Saimiri boliviensis boliviensis)

The squirrel monkey (Saimiri boliviensis boliviensis) has a well-defined breeding season during which adult males undergo androgen-dependent morphological changes, with acquisition of active spermatogenesis. To assess the hormonal events of this annual cycle, blood samples were obtained weekly from ten adult males, and serum was assayed for testosterone (T), androstenedione (ΔA), and dehydroepiandrosterone (DHEA). A significant seasonal variation was noted in mean serum T (P < 0.02), ΔA (P < 0.02), and DHEA (P < 0.001) concentrations. Mean ΔA concentrations increased from a nonbreeding season nadir of 91.4 ± 12.9 ng/ml (mean ± standard error) to a prebreeding concentration of 139 ± 10.5 ng/ml and breeding season peak of 167.5 ± 15.4 ng/ml (P < 0.05). Mean DHEA concentrations increased from a nonbreeding season nadir of 8.3 ± 0.8 to a breeding season peak of 14.3 ± 1.2 (P < 0.001). Mean T levels in the nonbreeding (52.2 ± 11.6 ng/ ml) and prebreeding season (48.6 ± 7.4) were similar. However, T significantly increased during the breeding season to 103.5 ± 12.8 ng/ml (P < 0.05). Progressive changes in body weight and morphology paralleled the rise in serum ΔA levels. The pattern of peripheral serum androgen concentrations throughout the year would suggest annual activation of the hypothalamic-pituitary-adrenal and/or hypothalamic-pituitary-gonadal axes.     

Presence of platelet-activating factor in squirrel monkey (Saimiri boliviensis) spermatozoa: seasonal differences

Platelet-activating factor (1-O-alkyl-2-acetyl-sn-glycero-3-phosphorylcholine) (PAF) is a potent signaling phospholipid which has pleiotropic biological properties in addition to platelet activation. PAF has been detected in the spermatozoa in a number of species. The concentration of PAF is inversely related to human spermatozoal quality. There are no reports on the presence of PAF in nonhuman primate spermatozoa. Therefore, the primary objective of this study was to determine if PAF is present in the spermatozoa from the squirrel monkey (which is a seasonal breeder). A second objective was to determine if PAF levels change from the breeding to the nonbreeding season. Endogenous lipids were extracted from mature Bolivian squirrel monkeys (Saimiri boliviensis) spermatozoa and assayed for the presence of PAF by [¹²⁵I] radioimmunoassay. PAF was detected in all samples assayed. PAF levels were significantly higher (P< 0.01) during the breeding season (mean: 3.58 ng/10⁶ spermatozoa) than the nonbreeding season (mean: 0.76 ng/10⁶ spermatozoa). The data demonstrate that PAF is present in squirrel monkey spermatozoa, with higher levels found during the breeding season. Additional studies are warranted to elucidate the role of PAF in spermatozoa function. Am. J. Primatol. 45:301–305, 1998. © 1998 Wiley-Liss, Inc.     

Seasonal weight changes in male rhesus monkeys (Macaca mulatta)

Adult male rhesus monkeys lose weight during the breeding season and regain it during the nonbreeding season. The annual pattern of maximum weight gain just prior to the onset of breeding resembles the seasonal “fattening” seen in squirrel monkeys, but the period of weight gain is less discrete. The magnitude of weight change is less in younger males, in that sexually immature males gain weight in both seasons, but significantly less during the breeding season. Females do not lose weight during the breeding season. Post hoc analyses revealed no significant correlations between male testosterone levels, dominance ranks, weights, or weight changes. The heaviest animals as juveniles were predictably the heaviest as adolescents. The timing of seasonal changes in testosterone did not correlate with the timing of changes in weight; weight losses followed the rise in testosterone, and weight gains continued until early in the breeding season after testosterone levels had already begun to rise. It is suggested that seasonal hormonal changes may influence activities in individuals and that changes in the activities of particular group members may alter the activity patterns of other group members. This alteration of activity patterns due to group influences on individuals as well as individual influences on the group may explain why hormonal regulation of seasonal weight appears to be indirect and why individuals (juveniles) experiencing no seasonal hormonal changes nonetheless show differences in activity patterns and seasonal weight changes.     

Low sex steroids, high steroid receptors: Increasing the sensitivity of the nonreproductive brain

Male aggressive behavior is generally regulated by testosterone (T). In most temperate breeding males, aggressive behavior is only expressed during the reproductive period. At this time circulating T concentrations, brain steroid receptors, and steroid metabolic enzymes are elevated in many species relative to the nonreproductive period. Many tropical birds, however, display aggressive behavior both during the breeding and the nonbreeding season, but plasma levels of T can remain low throughout the year and show little seasonal fluctuation. Studies on the year-round territorial spotted antbird (Hylophylax n. naevioides) suggest that T nevertheless regulates aggressive behavior in both the breeding and nonbreeding season. We hypothesize that to regulate aggressive behaviors during the nonbreeding season, when T is at its minimum, male spotted antbirds increase brain sensitivity to steroids. This can be achieved by locally up-regulating androgen receptors (ARs), estrogen receptors (ERs), or the enzyme aromatase (AROM) that converts T into estradiol. We therefore compared mRNA expression of AR, ERalpha, and AROM in free- living male spotted antbirds across reproductive and nonreproductive seasons in two brain regions known to regulate both reproductive and aggressive behaviors. mRNA expression of ERalpha in the preoptic area and AR in the nucleus taeniae were elevated in male spotted antbirds during the nonbreeding season when circulating T concentrations were low. This unusual seasonal receptor regulation may represent a means for the year-round regulation of vertebrate aggressive behavior via steroids by increasing the brain's sensitivity to sex steroids during the nonbreeding season.     

Immunoreactive inhibin levels in peripheral blood during the breeding season in the female Japanese monkey

Serum levels of immunoreactive inhibin as well as FSH, LH, estradiol-17 beta, and progesterone were measured by RIA in four mature female Japanese monkeys (Macaca fuscata fuscata) during the breeding season and subsequent transition into the nonbreeding season. During the breeding season, each monkey showed 2-6 ovulations, which were inferred from underlying endocrine events. The concentrations of serum inhibin increased during the luteal phase, but were low during the follicular phase. Such changes in serum inhibin levels correlated positively with those in serum progesterone levels. Basal levels of serum inhibin also increased during the breeding season, decreased during transition from the breeding season, and were low during the nonbreeding season. The parallel change in serum levels of inhibin and progesterone together with the increased basal levels of serum inhibin during this period suggests that both the CL and antral follicles are sources of circulating inhibin. Decreases in serum FSH levels during the luteal phase suggest that secretion of FSH is controlled by an inhibitory action of ovarian inhibin in addition to steroid hormones.     

Effect of age and breeding season on the developmental capacity of oocytes from unstimulated and follicle-stimulating hormone-stimulated rhesus monkeys

Effects of age and season on the developmental capacity of oocytes from unstimulated and FSH-stimulated rhesus monkeys were examined. Immature cumulus-oocyte complexes were matured in vitro in modified CMRL-1066 medium containing 20% bovine calf serum and subjected to in vitro fertilization followed by embryo culture. After fertilization, ova from unstimulated prepubertal monkeys displayed lower development to morula (4%) than those from unstimulated adult females (18% in breeding season and 22% in nonbreeding season). No developmental difference was found between ova from unstimulated adult monkeys in breeding and nonbreeding seasons. However, ova from FSH-primed prepubertal monkeys displayed greater development to blastocyst stage (54%) than those from adult monkeys in the breeding season (16%) and nonbreeding season (0%); and ova from FSH-primed adult females in the breeding season had significantly (P < 0.05) greater developmental competence than those obtained in the nonbreeding season (> or = morula stage, 54% vs. 3%; blastocyst stage, 16% vs. 0%). These data indicate that 1) rhesus monkey oocytes acquire developmental competence in a donor age-dependent manner, and 2) animal age and breeding season modulate the effect of FSH on oocyte developmental competence in the rhesus monkey.     

Physiological role for the cochaperone FKBP52 in androgen receptor signaling

Molecular chaperones mediate multiple aspects of steroid receptor function, but the physiological importance of most receptor-associated cochaperones has not been determined. To help fill this gap, we targeted for disruption the mouse gene for the 52-kDa FK506 binding protein, FKBP52, a 90-kDa heat shock protein (Hsp90)-binding immunophilin found in steroid receptor complexes. A mouse line lacking FKBP52 (52KO) was generated and characterized. Male 52KO mice have several defects in reproductive tissues consistent with androgen insensitivity; among these defects are ambiguous external genitalia and dysgenic prostate. FKBP52 and androgen receptor (AR) are coexpressed in prostate epithelial cells of wild-type mice. However, FKBP52 and AR are similarly coexpressed in testis even though testis morphology and spermatogenesis in 52KO males are usually normal. Molecular studies confirm that FKBP52 is a component of AR complexes, and cellular studies in yeast and human cell models demonstrate that FKBP52 can enhance AR-mediated transactivation. AR enhancement requires FKBP52 peptidylprolyl isomerase activity as well as Hsp90-binding ability, and enhancement probably relates to an affect of FKBP52 on AR-folding pathways. In the presence of FKBP52, but not other cochaperones, the function of a minimally active AR point mutant can be dramatically restored. We conclude that FKBP52 is an AR folding factor that has critically important physiological roles in some male reproductive tissues.     

Progesterone as a predictor of cyclicity in Bolivian squirrel monkeys during the breeding season

In a squirrel monkey breeding colony, two distinct groups of females were observed during the breeding season, December through March. One had low and the other had high estradiol (E2) and progesterone (P) concentrations. The conception rate in females with high E2 and P values was 74%. However, only 25% of monkeys with low steroid concentrations became pregnant during the breeding season. This study showed that all mature females in a colony may not be cycling concurrently and that two serum P measurements obtained at four-day intervals may be utilized to detect noncycling monkeys during the breeding season.     

Aggressive interactions differentially modulate local and systemic levels of corticosterone and DHEA in a wild songbird

During the nonbreeding season, when gonadal androgen synthesis is basal, recent evidence suggests that neurosteroids regulate the aggression of male song sparrows. In particular, dehydroepiandrosterone (DHEA) is rapidly converted in the brain to androgens in response to aggressive interactions. In other species, aggressive encounters increase systemic glucocorticoid levels. However, the relationship between aggression and local steroid levels is not well understood. Here, during the breeding and nonbreeding seasons, we tested the effects of a simulated territorial intrusion (STI) on DHEA and corticosterone levels in the brachial and jugular plasma. Jugular plasma is enriched with neurosteroids and provides an indirect index of brain steroid levels. Further, during the nonbreeding season, we directly measured steroid levels in the brain and peripheral tissues. Both breeding and nonbreeding males displayed robust aggressive responses to STI. During the breeding season, STI increased brachial and jugular corticosterone levels and jugular DHEA levels. During the nonbreeding season, STI did not affect plasma corticosterone levels, but increased jugular DHEA levels. During the nonbreeding season, STI did not affect brain levels of corticosterone or DHEA. However, STI did increase corticosterone and DHEA concentrations in the liver and corticosterone concentrations in the pectoral muscle. These data suggest that 1) aggressive social interactions affect neurosteroid levels in both seasons and 2) local steroid synthesis in peripheral tissues may mobilize energy reserves to fuel aggression in the nonbreeding season. Local steroid synthesis in brain, liver or muscle may serve to avoid the costs of systemic increases in corticosterone and testosterone.     

Testicular Expression of NGF,TrkA and p75 During Seasonal Spermatogenesis of the Wild Ground Squirrel (Citellus Dauricus Brandt)

The nerve growth factor (NGF) not only has an essential effect on the nervous system, but also plays an important role in a variety of non-neuronal systems, such as the reproductive system. The aim of this study was to compare the quality and quantity in expression of NGF and its receptors (TrkA and p75) in testes of the wild ground squirrel during the breeding and nonbreeding seasons. Immunolocalization for NGF was detected mainly in Leydig cells and Sertoli cells in testes of the breeding and nonbreeding seasons. The immunoreactivity of TrkA was highest in the elongated spermatids, whereas p75 in spermatogonia and spermatocytes in testes of the breeding season. In the nonbreeding season testes, TrkA showed positive immunostainings in Leydig cells, spermatogonia and primary spermatocytes, while p75 showed positive signals in spermatogonia and primary spermatocytes. Consistent with the immunohistochemical results, the mean mRNA and protein level of NGF and TrkA were higher in the testes of the breeding season than in non-breeding season, and then decreased to a relatively low level in the nonbreeding season. In addition, the concentration of plasma gonadotropins and testosterone were assayed by radioimmunoassay (RIA), and the results showed a significant difference between the breeding and nonbreeding seasons with higher concentrations in breeding season. In conclusion, these results of this study provide the first evidence on the potential involvement of NGF and its receptor, TrkA and p75 in the seasonal spermatogenesis and testicular function change of the wild ground squirrel.Key words: NGF, p75, seasonal spermatogenesis, TrkA, wild ground squirrel     

Conversion of androgens to estrogens in the male squirrel monkey (saimiri sciureas)

Many New World primates such as the squirrel monkey have extraordinarily high plasma levels of steroid hormones including cortisol, testosterone, progesterone and vitamin D₃. While plasma estrogen levels in female squirrel monkeys apparently are approximately the same as those found in other species no information is available for males. The present results indicate that the plasma levels of estrone (E1), estradiol (E2), and E1 sulfate are approximately 10-fold higher than those found in men. Comparative studies of androgen metabolism in genital skin fibroblasts indicate that squirrel monkey cells have higher aromatase and lower 5--reductase activity than human cells. Estimation of aromatase activity by a radiometric assay indicates that the high plasma estrogens are derived by peripheral conversion from testicular and/or adrenal androgens.     

A kidney epithelial cell line from a Bolivian squirrel monkey

Squirrel monkeys are the most commonly used New World primates in biomedical research, but in vitro studies are restricted by the limited number of cell lines available from this species. We report here the development and characterization of a continuous, kidney epithelial cell line (SQMK-FP cells) derived from a newborn squirrel monkey. Karyotype was consistent with Bolivian squirrel monkey (submetacentric chromosome pair 15 and acrocentric chromosome pair 16). All cells examined were hyperdiploid with chromosome numbers ranging from 52 to 57. Ultrastructural analysis of SQMK-FP cells revealed the presence of cell junctions with radiating filaments, indicating desmosomes and numerous surface projections containing longitudinally oriented filaments typical of tubular epithelium. Biochemically, SQMK-FP cells exhibit glucocorticoid resistance typical of the squirrel monkey. Glucocorticoid receptor (GR) binding is low in SQMK-FP cells because of high expression of the FK506-binding immunophilin FKBP51 that inhibits GR binding. SQMK-FP cells constitute a tubular epithelial cell line that has biochemical properties characteristic of squirrel monkeys and represents an alternate cell model to B-lymphoblast SML cells to study the biology of the squirrel monkey in vitro.     

Hormonal and behavioral changes at puberty in the squirrel monkey

Developmental changes in the reproductive behavior and physiology of 9 male and 15 female juvenile squirrel monkeys were evaluated in a 20-month study. Plasma levels of gonadal steroids remained relatively low for this species until most animals reached puberty between 2.5 and 3 years of age. Longitudinal assessment of plasma progesterone levels indicated that the onset of ovarian cycles tended to be synchronized between females although the 5 heaviest females began to cycle earlier than the rest. The heavier females reached puberty at a time which was appropriate to their birth in the wild, whereas most of the remaining females conceived 6 months later during a second period of reproductive activity that coincided with the laboratory mating season. Pubescent males underwent their first seasonal elevation in plasma testosterone levels during the second period and its onset was synchronized across all males. Thus, even in the absence of adults, pubertal processes in the squirrel monkey were strongly influenced by the seasonal breeding pattern. In addition, behavioral observations revealed that social maturation closely parallels reproductive ability in females, whereas males enter a protracted subadult stage after puberty.     

Seasonal changes in plasma testosterone and ejaculatory capacity in squirrel monkeys (Saimiri sciureus)

Seasonal changes in body weight, plasma testosterone and ejaculatory capacity were observed in five intact and two testosterone-implanted castrated squirrel monkeys for a total of 13 months. Electroejaculation was employed for obtaining data concerning ejaculation. In the intact animals, there were significant increases in body weight, ejaculate volume and plasma testosterone during the breeding season. With the exception of one animal, there was also a decrease in ejaculation latency during the season. Seasonal differences in the sperm count and sperm motility were not observed. Testosterone-implanted castrates showed changes in both ejaculate volume and ejaculation latency similar to those seen in intact monkeys during the breeding season. The body weight and plasma testosterone of the castrates remained quite constant throughout the year. Supported by NIH grants HD 00778, MH 21178 and MH 23645.